Amphiregulin is a heparin-binding glycoprotein of approximately 20 kDa which was originally purified from phorbol ester-treated MCF-7 human breast carcinoma cells. The factor belongs to the EGF-family of growth factors and stimulates the proliferation of several cell types (including keratinocytes and some fibroblast cell lines), while inhibiting the proliferation of other cells (including many carcinoma cell lines) (Shoyab et al., Proc. Natl. Acad. Sci. USA 85, 6528–6532 (1988); Shoyab et al., Science 243, 1074–1076 (1989)). Studies have revealed that AR is a major keratinocyte autocrine factor (Cook et al., Mol. Cell. Biol., 11, 2547–2557 (1991); Cook et al., In Vitro Cell. Dev. Biol. 28A, 218–222 (1992); Piepkorn et al., J. Cell. Physiol. 159, 114–120 (1994)). A special feature of AR is that its biological activity is completely blocked in the presence of heparin sulphate. AR has been reported to be the only growth factor displaying this property (Cook et al., Mol. Cell. Biol., 11, 2547–2557 (1991); Cook et al., In Vitro Cell. Dev. Biol. 28A, 218–222 (1992), Piepkorn et al., J. Cell. Physiol. 159, 114–120 (1994)). An isolated AR is disclosed in U.S. Pat. No. 5,115,096 (This and all other U.S. patents and patent applications cited herein are hereby incorporated by reference in their entirety).
Multiple soluble active isoforms of AR are produced by the proteolytic cleavage of a membrane-anchored AR precursor. These active isoforms are AR78, AR81, AR84, AR87, AR92, and AR98, etc. (Shoyab et al, Science 243, 1074–1076 (1989), Plowman et al., Mol. Cell. Biol. 10, 1969–1981 (1990); Thompson, S. A., et al., J. Biol. Chem. 271(30): 17927–31 (1996)). Soluble, secreted AR exerts its tyrosine phosphorylation and mitogenic affects largely through binding and activation of the 170 kDa EGFR (Johnson, G. R., et al., J. Biol. Chem. 268: 2924–2931 (1993)).
Immunolabeling and northern analyses have indicated that AR is low to undetectable in normal adult epidermis and markedly over-expressed in some neoplastic and non-neoplastic hyperproliferative disorders of the epidermis, including appendageal tumors, actinic keratoses, verrucae, and sequamous cell carcimomas (Piepkorn, et al., Am. J. Dermatol. 18: 165–171 (1996)). Studies have demonstrated that AR acts as an autocrine growth factor in human colon cancer. Basolateral administration of neutralizing antibodies against AR reduces the basal DNA replication of colon cancer cells (Damstrup, L. et al., Br. J. Cancer 80(7): 1012–9 (1999)).
It has been reported that mRNA expression of AR is dramatically elevated in psoriatic skin (Cook, et al., Cancer Res. 52: 3224–3227 (1992)). There is also strong AR staining in the cytoplasm of keratinocytes with psoriatic epidermis, in contrast to the sparse, focal labeling restricted to keratinocyte nuclei in normal skin. Keratinocyte-specific transgenic expression results in lesions with striking histological similarity to human psoriasis (Cook, et al., J. Clin. Invest. 100: 2286–2294 (1997)).
Published patent application EP 1249237A1 discloses an AR expression inhibitor and the use thereof as an ameliorating agent for aged skin.
International patent application WO 90/14069 and U.S. Pat. No. 5,262,298 disclose the isolation of keratineocyte autocrine factor (KAF) having the matched amino acid sequence of AR. These two references generally discuss that administration of an inhibitor of KAF activity would benefit conditions such as psoriasis as well as squamous cell carcinomas.
U.S. Pat. No. 5,830,995 and U.S. Pat. No. 6,204,359 disclosed antibodies against AR.
The present invention is directed to neutralizing anti-AR antibodies having the disclosed amino acid sequences or binding characteristics, which are not disclosed in the above-referenced publications. The in vivo efficacy of the anti-AR antibodies for treating psoriasis and cancer are tested in the present invention. These antibodies will find use for inhibiting cancer cell growth, wound healing, enhancing skin quality, or/and treating psoriasis in a subject. The present invention is also pursuant to methods of treating psoriasis or epidermal and pancreatic cancers with any antagonists of AR, preferably, anti-AR antibodies, and more preferably, the antibodies claimed herein.